RESUMEN
Vitiligo is a chronic autoimmune disease characterized by the T helper 1 (Th1) cytokine-driven immune destruction of melanocytes (MCs). Although narrowband ultraviolet B (NBUVB) phototherapy has been proven to be an effective therapeutic option, the repigmentation response to that phototherapy varies greatly in different vitiligo patients. Here, we demonstrate that there is an increase of NBUVB-induced cellular senescence in vitiligo MCs exposed to Th1 cytokine interferon γ (IFNγ) and/or tumor necrosis factor α (TNFα) in lesional vitiligo skin from poor responders who had undergone NBUVB phototherapy. Supplementation with exogenous recombinant human stem cell factor (rhSCF) in the culture medium as well as the lentiviral vector-mediated overexpression of cKIT could prevent the MCs from the IFNγ/TNFα-accelerated cellular senescence. Mechanistic studies indicated that the reduced ratio of membrane-bound KIT (mKIT) to the soluble form of KIT (sKIT) is directly related to the cellular senescence of vitiligo MCs following exposure to IFNγ and TNFα. Furthermore, the matrix metalloprotease 9 (MMP9) inhibitor GM6001 attenuates the production of sKIT via the suppression of cKIT ectodomain shedding. Altogether, our study indicates that the presence of Th1 cytokines IFNγ and/or TNFα in the epidermal milieu might impair the repigmentation response of vitiligo patients to NBUVB phototherapy.
Asunto(s)
Vitíligo , Humanos , Vitíligo/radioterapia , Vitíligo/tratamiento farmacológico , Factor de Necrosis Tumoral alfa , Citocinas , Interferón gamma , Fototerapia , Melanocitos/patología , Resultado del Tratamiento , AceleraciónRESUMEN
BACKGROUND: Few studies have addressed the impact of the psoriasis-related proinflammatory cytokines on the proliferation and melanogenesis of melanocytes (MCs) in lesional psoriatic skin. OBJECTIVE: We investigated the effects of TNFα, IL17A, and IL8 on the proliferation and melanin synthesis of MCs. METHODS: Skin specimens were biopsied from patients with psoriasis vulgaris at the active stage, or from the tail skin of Dct-LacZ mice with imiquimod (IMQ)-induced psoriasiform dermatitis. Cultured keratinocytes (KCs), MCs, and human skin explants were used in this study. The numbers of MCs were measured via ß-galactosidase staining, EdU incorporation and HMB45 immunohistochemical staining. The expression of human ß-defensin 3 (hBD3) in KCs was silenced by siRNA, the conditioned medium (CM) from siRNA-transfected KCs was used to treat MCs, then followed by αMSH stimulation. The melanogenesis-related genes were examined by using qRT-PCR and western blotting. RESULTS: The increased number of MCs and decreased melanin content were highly relevant to the enhanced expression of IL8 and BD3 both in human psoriatic skin and in IMQ-treated mouse tail skin. IL8 expression in KCs and CXCR2 expression in MCs was significantly increased by IL17A and TNFα, the αMSH-induced upregulations of microphthalmia-associated transcription factor (MITF) and tyrosinase in MCs were abrogated by the CM from hBD3-unsilenced KCs, but not from hBD3-silenced KCs. CONCLUSION: Our results suggest the roles of IL8-CXCR2 activation in promoting MC proliferation and of BD3 upregulation in reducing melanogenesis. These findings have been implicated in the underlying mechanism that active psoriasis prefers hypopigmentation despite chronic inflammation.
Asunto(s)
Psoriasis , Factor de Necrosis Tumoral alfa , Humanos , Animales , Ratones , Factor de Necrosis Tumoral alfa/metabolismo , Melaninas/metabolismo , Citocinas/metabolismo , Interleucina-8/metabolismo , Melanocitos/metabolismo , Psoriasis/metabolismo , ARN Interferente Pequeño/metabolismo , Imiquimod/farmacología , Proliferación Celular , Factor de Transcripción Asociado a Microftalmía/metabolismoRESUMEN
Long noncoding RNAs (lncRNAs) are the most recently discovered class of noncoding RNAs. LncRNAs play a crucial role in multiple disorders. However, the role and mechanism of action of lncRNAs in keloids remain unclear. Here, qRT-PCR and western blotting assays were performed to determine the expression of genes and proteins, respectively. MTT assays were carried out to measure the proliferation of keloid fibroblasts. In addition, a luciferase activity assay was conducted to investigate the relationships between LINC00937 and miR-28-5p and between miR-28-5p and MC1R. The results showed that LINC00937 and MC1R were decreased, whereas miR-28-5p was increased in keloid tissues. LINC00937 overexpression in keloid fibroblasts could repress the extracellular matrix (ECM) deposition and cell proliferation and promote MC1R expression. Moreover, high expression of miR-28-5p and low expression of LINC00937 were detected in keloid fibroblasts. We further showed that LINC00937 promoted MC1R expression by sponging miR-28-5p. Finally, our data indicated that LINC00937 inhibited the ECM deposition and proliferation of keloid fibroblasts by inhibiting miR-28-5p and facilitating MC1R expression. Overall, LINC00937 suppressed the ECM deposition and proliferation of keloid fibroblasts by acting as an miR-28-5p sponge and promoting MC1R expression. Our data suggested that LINC00937 is a potential target for keloid treatment.
Asunto(s)
Proliferación Celular , Matriz Extracelular/metabolismo , Fibroblastos/metabolismo , Queloide/metabolismo , MicroARNs/metabolismo , ARN Largo no Codificante/metabolismo , Receptor de Melanocortina Tipo 1/metabolismo , Piel/metabolismo , Adolescente , Adulto , Células Cultivadas , Matriz Extracelular/genética , Matriz Extracelular/patología , Femenino , Fibroblastos/patología , Humanos , Queloide/genética , Queloide/patología , Masculino , MicroARNs/genética , Persona de Mediana Edad , ARN Largo no Codificante/genética , Receptor de Melanocortina Tipo 1/genética , Transducción de Señal , Piel/patología , Adulto JovenRESUMEN
Although vitamin C (VC, L-ascorbic acid) has been widely used as a skin lightening agent for a long time, the mechanism by which it inhibits melanogenesis remains poorly understood. It is well-documented that the intramelanocytic pH is an important factor in regulating tyrosinase function and melanosome maturation. The activity of tyrosinase, the rate-limiting enzyme required for melanin synthesis, is generally minimal in an acidic environment. Given that VC is an acidic compound, we might speculate that the intracellular acidification of melanocytes induced by VC likely reduces melanin content through the suppression of tyrosinase activity. The results of this study reveal that treatment of melanocytes with VC or its derivatives, magnesium ascorbyl phosphate (MAP) and 3-O-ethyl-L-ascorbic acid (AAE), resulted in significant decreases in the tyrosinase activity and melanin content and in the levels of intracellular reactive oxygen species (ROS), indicating that VC and its derivatives possess antimelanogenic and antioxidative activities. Western blotting analysis indicated that VC, MAP, and AAE exert their antimelanogenic activity by inhibiting the tyrosinase activity rather than by downregulating the expression of melanogenic proteins such as tyrosinase, premelanosome protein 17 (Pmel17) and microphthalmia-associated transcription factor (MITF). Further, we found that the reduced tyrosinase activity of melanocytes treated with VC or its derivatives could be reactivated following intracellular neutralization induced by ammonium chloride (NH4Cl) or concanamycin A (Con A). Finally, we examined the expression of sodium-dependent VC transporter-2 (SVCT-2) using western blotting and qPCR, which revealed that there was a significant increase in the expression of SVCT-2 in melanocytes following treatment with VC. VC-mediated intracellular acidification was neutralized by phloretin (a putative SVCT-2 inhibitor) in a dose-dependent manner. Taken together, these data show that VC and its derivatives suppress tyrosinase activity through cytoplasmic acidification that potentially results from enhanced VC transmembrane transport via the VC transporter SVCT-2.
Asunto(s)
Antioxidantes/metabolismo , Ácido Ascórbico/metabolismo , Citoplasma/metabolismo , Melaninas/metabolismo , Melanocitos/fisiología , Melanosomas/metabolismo , Monofenol Monooxigenasa/metabolismo , Preparaciones para Aclaramiento de la Piel/metabolismo , Animales , Ácido Ascórbico/análogos & derivados , Diferenciación Celular , Línea Celular , Citoplasma/química , Humanos , Concentración de Iones de Hidrógeno , Ratones , Ratones Endogámicos C57BL , Pigmentación de la Piel , Transportadores de Sodio Acoplados a la Vitamina C/genética , Transportadores de Sodio Acoplados a la Vitamina C/metabolismo , Regulación hacia ArribaRESUMEN
Excessive expansion of the transit-amplifying (TA) cell compartment is a distinct morphological characteristic of psoriatic epidermal hyperplasia. In order to examine the activation of basal stem cells and how they replenish such an enlarged compartment of TA cells in psoriatic epidermis, we utilized a BrdU labeling method to monitor mitotic stem cells in a mouse model of psoriasiform dermatitis, which was induced by imiquimod. Our results showed that perpendicular and parallel cell division characteristics of dividing stem cells existed in the inflamed epidermis. When we analyzed templateDNA strand segregation in trypsin-dissociated human psoriatic keratinocytes using BrdU pulse-chase labeling, we found that the percentage of asymmetric segregation of BrdU was significantly increased in the cell pairs of psoriatic epidermal cells compared with normal epidermal cells. Furthermore, we also examined the effects of both interleukin (IL)-17A and IL-22 cytokines on the differentiation status of cultured human keratinocytes. The results indicated that both cytokines had synergistic effects on passage-one epidermal cell sheets derived from skin explants and also on cultured keratinocytes, were involved in the maintenance of the undifferentiated stem cell phenotype, and these results suggest an efficient mechanism for preventing the premature loss of basal stem-cell pools in the pro-inflammatory cytokine-enriched milieu of the psoriatic epidermis. Our findings suggest that inhibition of hyperactive stem cells represents a potential therapeutic target to combat recalcitrant epidermal hyperplasia in psoriasis.
Asunto(s)
Diferenciación Celular/genética , Interleucina-17/genética , Interleucinas/genética , Psoriasis/genética , Animales , División Celular Asimétrica/genética , Modelos Animales de Enfermedad , Epidermis/metabolismo , Humanos , Interleucina-17/metabolismo , Interleucinas/metabolismo , Queratinocitos/metabolismo , Queratinocitos/patología , Ratones , Psoriasis/metabolismo , Psoriasis/patología , Células Madre/citología , Células Madre/metabolismo , Interleucina-22RESUMEN
Intricate coordinated mechanisms that govern the synchrony of hair growth and melanin synthesis remain largely unclear. These two events can be uncoupled in prematurely gray hair, probably due to oxidative insults that lead to the death of oxidative stress-sensitive melanocytes. In this study, we examined the gene expression profiles of middle (bulge) and lower (hair bulb) segments that had been micro-dissected from unpigmented and from normally pigmented hair follicles from the same donors using quantitative real-time RT-PCR (qPCR) arrays. We found a significant down-regulation of melanogenesis-related genes (TYR, TYRP1, MITF, PAX3, POMC) in unpigmented hair bulbs and of marker genes typical for melanocyte precursor cells (PAX3, SOX10, DCT) in unpigmented mid-segments compared with their pigmented analogues. qPCR, western blotting and spin trapping assays revealed that catalase protein expression and hydroxyl radical scavenging activities are strongly repressed in unpigmented hair follicles. These data provide the first clear evidence that compromised antioxidant activity in gray hair follicles simultaneously affects mature hair bulb melanocytes and their immature precursor cells in the bulge region.
Asunto(s)
Antioxidantes/metabolismo , Color del Cabello , Cabello/metabolismo , Melanocitos/metabolismo , Adulto , Secuencia de Bases , Western Blotting , Cartilla de ADN , Femenino , Perfilación de la Expresión Génica , Humanos , Masculino , Reacción en Cadena de la Polimerasa , Marcadores de Spin , Adulto JovenRESUMEN
Activation of the α-melanocyte-stimulating hormone (αMSH)/melanocortin-1 receptor (MC1R) signalling pathway exerts antagonistic actions on cutaneous inflammatory and fibrogenic responses in addition to promoting pigment production. Herein, the expression of MC1R by keloid-derived fibroblasts and keloid scar tissue was investigated using a range of techniques. MC1R mRNA expression levels in five different keloid fibroblast cell lines were significantly reduced to less than half compared with five normal fibroblast cell lines (P < 0.05). Immunohistological analysis of tissue samples indicated that MCR1 immunoreactivity in both epidermal and dermal compartments of five keloid tissue samples was dramatically decreased compared with normal skin (P < 0.05). Insufficient expression of MC1R on human dermal fibroblasts might abolish the αMSH-mediated suppression of collagen production and myofibroblast transformation elicited by the profibrotic cytokine-transforming growth factor-ß1. Restoration of reduced MC1R by dermal fibroblasts may lead to novel scar-reducing therapeutic approaches for treating this refractory fibrotic disease.